No comparative study of the effects of sphingolipid metabolites on proliferation and differentiation in normal human breast epithelial cells versus stem cells and tumorigenic cells has been reported. The purpose of this study was to evaluate the chemotherapeutic and chemopreventive potential of sphingoid bases (sphingosine and sphinganine) using a novel cell culture system of normal human breast epithelial cells (HBEC) developed from breast tissues of healthy women obtained during reduction mammoplasty (Type I HBEC with stem cell characteristics and Type II HBEC with basal epithelial cell phenotypes) and transformed tumorigenic Type I HBEC. The results show that sphinganine inhibited the growth and induced apoptosis of transformed tumorigenic Type I HBEC more potently than sphingosine (IC50 for sphinganine 4 μM; sphingosine 6.4 μM). Both sphinganine and sphingosine at high concentrations (8–10 μM) arrested the cell cycle at G2/M. Sphinganine inhibited the growth and caused death of Type I HBEC more strongly than sphingosine. In comparison, Type II HBEC (normal differentiated cells) were less sensitive to the growth-inhibitory effects of sphingoid bases than Type I HBEC (stem cells) or transformed tumorigenic Type I HBEC, suggesting that sphingoid bases may serve as chemotherapeutic agents. At concentrations (0.05, 0.1, and 0.5 μM) that are below the growth-inhibitory range, sphingoid bases induced differentiation of Type I HBEC to Type II HBEC, as detected morphologically and via expression of a tumor suppressor protein, maspin, which is a marker of Type II HBEC. Thus, sphingoid bases may function as chemotherapeutic as well as chemopreventive agents by preferentially inhibiting cancer cells and eliminating stem cells from which most breast cancer cells arise.
American Cancer Society.Cancer Facts and Figures. Atlanta, GA: American Cancer Society, 1999.
2.
JemalA, TiwariRC, MurrayT, GhafoorA, SamuelsA, WardE, FeuerEJ, ThunMJ. Cancer statistics. CA Cancer J. Clin54: 8–29, 2004.
3.
AhnEH, SchroederJJ. Bioactive sphingolipids are constituents of milk and soy products. J Food Sci67: 522–524, 2002.
4.
VesperH, SchmelzEM, Nikolova-KarakashianMN, DillehayDL, LynchDV, MerrillAHJr.Sphingolipids in food and the emerging importance of sphingolipids to nutrition. J Nutr129: 1239–1250, 1999.
5.
NilssonA.Metabolism of sphingomyelin in the intestinal tract of the rat. Biochim Biophys Acta164: 575–584, 1968.
6.
SchmelzEM, CrallKJ, LarocqueR, DillehayDL, MerrillAHJr.Uptake and metabolism of sphingolipids in isolated intestinal loops. J Nutr124: 702–712, 1994.
7.
NybergL, NilsonA, LundgrenP, DuanR-D.Localization and capacity of sphingomyelin digestion in the rat intestinal tract. J Nutr Biochem8: 112–118, 1997.
8.
ImaizumiK, TominagaA, SatoM, SuganoM.Effects of dietary sphingolipids on levels of serum and liver lipids in rats. Nutr Res12: 543–548, 1992.
9.
WangE, RossPF, WilsonTM, RileyRT, MerrillAHJr.Increases in serum sphingosine and sphinganine and decreases in complex sphingolipids in ponies given feed containing fumonisins, mycotoxins produced by fusarium moniliforme. J Nutr122: 1706–1716, 1992.
10.
AbnetCC, BorkowfCB, QiaoY-L, AlbertPS, WangE, MerrillAHJr., MarkSD, DongZ-W, TaylorPR, DawseySM. A cross-sectional study of human serum sphingolipids, diet and physiologic parameters. J Nutr131: 2748–2752, 2001.
11.
AhnEH, SchroederJJ. Sphingoid bases and ceramide induce apoptosis in HT-29 and HCT-116 human colon cancer cells. Exp Biol Med227: 345–353, 2002.
12.
SweeneyEA, SakakuraC, ShirahamaT, MasamuneA, OhtaH, HakomoriS-I, YasuyukiJ.Sphingosine and its methylated derivative N,N-dimethlysphingosine (DMS) induce apoptosis in a variety of human cancer cell lines. Int J Cancer66: 358–366, 1996.
13.
CaiZ, BettaiebA, MahdaniNE, LegresLG, StancouR, MaslliahJ, ChouaibS.Alteration of the sphingomylein/ceramide pathway is associated with resistance of human breast carcinoma MCF7 cells to tumor necrosis factor-mediated cytotoxicity. J Biol Chem272: 6918–6926, 1997.
14.
GillZP, PerksCM, NewcombPV, HollyJM. Insulin-like growth factor-binding protein (IGFBP-3) predisposes breast cancer cells to programmed cell death in a non-IGF-dependent manner. J Biol Chem272: 25602–25607, 1997.
15.
DillehayDL, WebbSK, SchmelzEM, MerrillAHJr.Dietary sphingomyelin inhibits 1,2-dimethylhydrazine-induced colon cancer in CF1 mice. J Nutr124: 615–620, 1994.
16.
SchmelzEM, DillehayDL, WebbSK, ReiterA, AdamsJ, MerrillAHJr.Sphingomyelin consumption suppresses aberrant colonic crypt foci and increases the proportion of adenomas versus adenocarcinomas in CF1 mice treated with 1,2-dimethylhydrazine: implications for dietary sphingolipids and colon carcinogenesis. Cancer Res56: 4936–4941, 1996.
17.
SchmelzEM, BushnevAS, DillehayDL, LiottaDC, MerrillAHJr.Suppression of aberrant colonic crypt foci by synthetic sphingomyelins with saturated or unsaturated sphingoid base backbones. Nutr Cancer28: 81–85, 1997.
18.
SchmelzEM, SullardsMS, DillehayDL, MerrillAHJr.Colonic cell proliferation and aberrant crypt formation are inhibited by dairy glycosphingolipids in 1,2-dimethylhydrazine-treated CF1 mice. J Nutr130: 522–527, 2000.
19.
LemonnierLA, DillehayDL, VespremiMJ, AbramsJ, BrodyE, SchmelzEM. Sphingomyelin in the suppression of colon tumors: prevention versus intervention. Arch Biochem Biophys419: 129–138, 2003.
20.
SymolonH, SchmelzEM, DillehayDL, MerrillAHJr.Dietary soy sphingolipids suppress tumorigenesis and gene expression in 1,2-dimethylhydrazine-treated CF1 mice and APCMin/+ mice. J Nutr134: 1157–1161, 2004.
21.
SchmelzEM, RobertsPC, KustinEM, LemonnierLA, SullardsMC, DillehayDL, MerrillAHJr.Modulation of intracellular β-catenin localization and intestinal tumorigenesis in vivo and in vitro by sphingolipids. Cancer Res61: 6723–6729, 2001.
22.
SakakuraC, SweeneyEA, ShirahamaT, HagiwaraA, YamaguchiT, TakahashiT, HakomoriS, IgarashiY.Selectivity of sphingosine-induced apoptosis: lack of activity of DL-erythro-dihydrosphingosine. Biochem Biophys Res Commun246: 827–830, 1998.
23.
BaekM-Y, YooH-S, NakayaK, MoonD-C, LeeY-M.Sphingolipid metabolic changes during chiral C2-ceramides induced apoptosis in human leukemia cells. Arch Pharm Res24: 144–149, 2001.
24.
GoswamiR, DawsonG.Does ceramide play a role in neuronal cell apoptosis?J Neurosci Res60: 141–149, 2000.
25.
RadinNS. Apoptotic death by ceramide: will the real killer please stand up?Med Hypotheses57: 96–100, 2000.
26.
KaoCY, NomataK, OakleyCS, WelschCW, ChangCC. Two types of normal human breast epithelial cells derived from reduction mammoplasty: phenotypic characterization and responses to SV 40 transfection. Carcinogenesis16: 531–538, 1995.
27.
ChangCC, SunW, CruzA, SatohM, TaiM-H, TroskoJE. A human breast epithelial cell type with stem cell characteristics as target cells for carcinogenesis. Radiat Res155: 201–207, 2001.
28.
KangK-S, MoritaI, CruzA, JeonYJ, TroskoJE, ChangCC. Expression of estrogen receptors in a normal human breast epithelial cell type with luminal and stem cell characteristics and its neoplastically transformed cell lines. Carcinogenesis18: 251–257, 1997.
29.
KaoCY, OakleyCS, WelschCW, ChangCC. Growth requirements and neoplastic transformation of two types of normal human breast epithelial cells derived from reduction mammoplasty. In Vitro Cell Dev Biol Anim33: 282–288, 1997.
30.
KangKS, SunW, NomataK, MoritaI, CruzA, LiuCJ, TroskoJE, ChangCC. Involvement of tyrosine phosphorylation of p185 (c-erbB2/neu) in tumorigenicity induced by x-rays and the neu oncogene in human breast ephithelial cells. Mol Carcinog21: 225–233, 1998.
31.
SunW, KangKS, MoritaI, TroskoJE, ChangCC. High susceptibility of a human breast epithelial cell type with stem cell characteristics to telomerase activation and immortalization. Cancer Res59: 6118–6123, 1999.
32.
CrawfordKW, BittmanR, ChunJ, ByunHS, BowenWD. Novel ceramide analogues display selective cytotoxicity in drug-resistant breast tumor cell lines compared to normal breast epithelial cells. Cell Mol Biol49: 1017–1023, 2003.
33.
SugawaraT, KinoshitaM, OhnishiM, NagataJ.Digestion of maize sphingolipids in rats and uptake of sphingadienine by Caco-2 cells. J Nutr133: 2777–2782, 2003.
34.
FrankfurtOS, SugarbarkerEV, RobbJA, VillaL.Synergistic induction of apoptosis in breast cancer cells by tamoxifen and calmodulin inhibitors. Cancer Lett97: 149–154, 1995.
35.
PerryRR, KangY, GreavesBS. Effects of tamoxifen on growth and apoptosis of estrogen-dependent and -independent human breast cancer cells. Ann Surg Oncol2: 238–245, 1995.
36.
GelmannEP. Tamoxifen induction of apoptosis in estrogen receptor-negative cancers: new tricks for an old dog?J Natl Cancer Inst88: 224–226, 1996.
37.
MandlekarS, YuR, TanTH, KongAN. Activation of caspase-3 and c-Jun NH2-terminal kinase-1 signaling pathways in tamoxifen-induced apoptosis of human breast cancer cells. Cancer Res60: 5995–6000, 2000.
38.
SenchenkovA, LitvakDA, CabotMC. Targeting ceramide metabolism—a strategy for overcoming drug resistance. J Natl Cancer Inst93: 347–357, 2001.
39.
ChangCC. Recent translational research: stem cells as the root of breast cancer. Breast Cancer Res8: 103 (doi:10.1186/bcr1385), 2006.
40.
OkazakiT, BellRM, HannunYA. Sphingomyelin turnover induced by vitamin D3 in HL-60 cells. J Biol Chem264: 19076–19080, 1989.
41.
OkazakiT, BielawskaA, BellRM, HannunYA. Role of ceramide as a lipid mediator of 1α, 25-dihydroxy vitamin D3-induced HL-60 cell differentiation. J Biol Chem265: 15823–15831, 1990.
42.
KimMY, LinardicC, ObeidL, HannunY.Identification of sphingomyelin turnover as an effector mechanism for the action of tumor necrosis factor α and γ–interferon-specific role in differentiation. J Biol Chem266: 484–489, 1991.
43.
TomanRE, SpiegelS, FadenAI. Role of ceramide in neuronal cell death and differentiation. J Neurotrauma17: 891–898, 2000.
44.
HuwilerA, KolterT, PfeilschifterJ, SandhoffK.Physiology and pathophysiology of sphingolipid metabolism and signaling. Biochim Biophys Acta1485: 63–99, 2000.
45.
BuccolieroR, FutermanAH. The roles of ceramide and complex sphingolipids in neuronal cell function. Pharm Res47: 409–419, 2003.
46.
HuiEKW, YangYH, YungY.Schedule-dependent sphinganine potentiation of retinoic acid-induced differentiation, cell-growth inhibition, and nucleophosmin translocation in a human leukemia-cell line (HL-60). Exp Hematol20: 454–461, 1992.
47.
SaitohM.Maspin, a tumor suppressor gene, is expressed in human basal breast epithelial cells but not in breast epithelial stem cells and breast cancer cells. MS thesis, Michigan State University, 2004.
48.
ParkJS, NohDY, KimSH, KimSH, KongG, ChangCC, LeeYS, TroskoJE, KangKS. Gene expression analysis in SV40-immortalized human breast luminal epithelial cells with stem cell characteristics using a cDNA microarray. Int J Oncol24: 1545–1558, 2004.
49.
MaassN, NagasakiK, ZiebartM, MundhenkeC, JonatW.Expression and regulation of tumor suppressor gene maspin in breast cancer. Clin Breast Cancer3: 281–287, 2002.
50.
ShiHY, LiangR, TempletonNS, ZhangM.Inhibition of breast tumor progression by systemic delivery of the maspin gene in a syngeneic tumor model. Mol Ther5: 755–761, 2002.
51.
ShiHY, ZhangW, LiangR, KittrellF, TempletonNS, MedinaD, ZhangM.Modeling human breast cancer metastasis in mice: maspin as a paradigm. Histol Histopathol18: 201–206, 2003.
52.
TaiM-H, ChangCC, OlsonLK, TroskoJE. Oct4 expression in adult human stem cells: evidence in support of the stem cell theory of carcinogenesis. Carcinogenesis26: 495–502, 2005.
